THC drug testing in the workplace and roadside - relevant intersection of science, society, and law.
https://www.youtube.com/watch?v=2f_0EwZ8Fk4
Saturday's FrankTalk: Fear and Impairment
THC drug testing in the workplace and roadside - relevant intersection of science, society, and law. https://www.youtube.com/watch?v=2f_0EwZ8Fk4
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A FrankTalk on cannabis DUI laws, workplace drug testing, and the current state of things.
Cost: Free event, open to the community Time: 12:00 PM to 04:00 PM, talk begins at 01:00. Location: Herman's Hideaway: 1578 S Broadway, Denver, CO 80210 Join Frank Conrad, Lab Director of Colorado Green Lab for The SCIENCE! of Cannabis. Doors at noon. Frank will talk from 1-1:45 and then open the floor for questions for 30 minutes. This is our first event on a Saturday! As always, we'll be there after the talk for a few hours or more having a few beers at the bar absolutely ready and thrilled to talk SCIENCE! If you can't make the talk, come for the hanging out after. The patio is nice and the weather has been beautiful. There is always a great mix of industry, scientists, cannabis patients, recreational users and curious non users. https://www.facebook.com/events/1817254101844140/ Another great FrankTalk last night, this one on the subject of cannabis terpenes and their role as antifungal compounds against powdery mildew and as insect deterrents. I'll post a link to the YouTube video of the talk once it's uploaded.
Thank you, World Viral, for recording and post-processing, and Herman's Hideaway for providing the venue! Video of Monday's FrankTalk: The Science! of Cannabis Cannabidiol Conversion to THC 15-August-2016dule Zero is real. https://www.facebook.com/events/1046200882102388/
Conversion of Cannabidiol (CBD) to Tetrahydrocannabinol The spontaneous conversion of CBD to THC has significant medical and legal implications. I'll be presenting the current evidence for this phenomenon at next week's FrankTalk, and how it may impact patients, care providers, growers, extractors, and manufacturers, as well as their legal counsel. Cost: Free event, open to the community This month, join us at a new location! Herman's Hideaway: 1578 S Broadway, Denver, CO 80210 Join Frank Conrad, Lab Director of Colorado Green Lab for The SCIENCE! of Cannabis. Frank will talk from 6:30-7:00 and then open the floor for questions. As always, we'll be there at 5:30 having a few beers at the bar if you want to come early to ask questions, hang out and have a drink. Even if you can't make the talk, come for the hanging out after. The patio is nice and the evening weather is beautiful. There is always a great mix of industry, scientists, cannabis patients, recreational users and curious non-users. August FrankTalk
The topic of August's FrankTalk (date and venue TBD) will be the conversion of CBD to THC, and the legal and medical implications of this little-known phenomenon. I'll update blog with more details and source references in the coming week, but for now, here's the summary: tl; dr Cannabidiol (CBD) and tetrahydrocannabinol (THC) are nearly identical, but whereas THC is psychoactive, CBD is not. The two compounds differ only slightly in chemical structure - CBD is bicyclic ("two-ring") and THC is tricyclic ("three-ring"). The carbon and oxygen molecules in the partial central ring of CBD will rearrange under various natural and artificial conditions to form THC. The rate of conversion is influenced by many factors, including length of storage time, exposure to light and air, ambient temperature, acidity, and the presence of protic solvents such as ethanol or methanol (alcohols). http://www.9news.com/news/local/hugo-water-tests-negative-for-thc/280096515
The Colorado Bureau of Investigation (CBI) has ruled out the presence of THC [1] in the water supply of Hugo, CO [2], but the identity of the contaminating compound has not yet been resolved or disclosed. I have a few thoughts though, which I will post later today. Side note: Incident Management Due to the nature of the suspected contaminant (the unlikelihood of THC), there has been some tacit criticism on social media about how this situation was handled by local officials. As mentioned in a previous post, the Duquenois–Levine test creates spurious results, but this is also not widely known. In my opinion, the Hugo water incident was handled very effectively, and with the utmost regard for the well-being of the town's residents. In just 48 hours:
If you require an example of how water contamination issues should *not* be handled, please read about Flint, MI [3]. In Hugo, the coordinated efforts of the Department of Public Works, Dr. John Fox of the Lincoln Community Hospital and Care Center, and Captain Michael Yowell of Lincoln County Sheriff's Office, were nothing short of commendable. References 1. Healy, J., A Colorado Town Tests Positive for Marijuana (in Its Water), in The New York Times. 2016. 2. Villanueva, R., Hugo water tests negative for THC. 2016, Channel 9 News, KUSA. 3. Graham, D.A., What Did the Governor Know About Flint's Water, and When Did He Know It?, in The Atlantic. 2016. Water Contamination in Hugo, Colorado On 21-July-2016, Officials in Hugo, Colorado reported that they had detected Δ9-tetrahydrocannabinol (THC), the psychoactive compound derived from Cannabis sativa [1, 2], in the town's water supply, and have issued a health advisory warning residents not to bathe or drink municipal water until further notice. Legal and Pharmacological Threshold of THC While there is no regulatory limit for THC in water that I'm aware of, there is a widely accepted threshold for THC in industrial hemp-derived products intended for human consumption. Products containing no more than 0.3% THC w/w (3000 ppm) are legal (even by the Federal government) and not considered psychoactive. By application of the same standard, 3000 ppm or lower in drinking water would produce no clinically relevant symptoms or noticeable effects. Solubility and Stability of THC THC is a non-polar compound (i.e. fat-soluble) and does not dissolve or mix well with water [1]. It is also susceptible to degradation by various environmental factors, and upon exposure to light, oxygen, or elevated ambient temperature, rapidly breaks down into inert compounds [3]. In other words, THC would not travel far in groundwater, and wouldn't survive very long even if it did. The Duquenois–Levine Marijuana Field Test The water supply in Hugo was evaluated using a color-change assay called the modified Duquenois–Levine Marijuana Field Test (DL test). As the name implies, the DL test is used to screen suspected contraband for the presence of THC and other cannabinoids. Despite its frequent use as a screening tool, it is not widely known that the modified Duquenois–Levine (DL) test reacts with numerous non-cannabinoid compounds [4-8]. For example, the DL test will produce a false positive THC result if used to test sage, cypress, or eucalyptus plant material [5, 6, 9] or over-the-counter sore throat lozenges [5, 6]. How It Works - Duquenois–Levine Marijuana Field Test (DL test) How It Works
In theory, if the DL field test kit produces two distinct color layers at the top and bottom of the sample vial - light blue, dark blue, or light purple layer (top) covering a dark purple layer (bottom) - then THC is present in the sample. In reality, many compounds (hexylresorcinol found in throat lozenges is just one example of something that produces false positives) will produce identical or similar color results with no THC present whatsoever [11]. The Duquenois component of the test reacts with resorcinol structures (found on both THC annd hexylresorcinol - see figure below), and the Levine modification allows fat-soluble molecules to pass into the chloroform layer at the bottom of the vial (both THC and hexylresorcinol are fat soluble). While it is unclear what contamination is actually in the Hugo water supply, it is unlikely to be THC, based on the spurious nature of the DL test and the poor water solubility of THC itself. Further testing is underway to resolve the question and results should be released tomorrow by the Department of Health. References
1. Tetrahydrocannabinol, in Wikipedia. 2016, Wikipedia. 2. Cannabis sativa, in Wikipedia. 2016. 3. Trofin I, D.G., Vaireanu D, Filipescu L, The Influence of Long-term Storage Conditions on the Stability of Cannabinoids derived from Cannabis Resin. Revista de Chimie (Bucharest), 2012. 63(4): p. 422-427. 4. Duquenois–Levine reagent, in Wikipedia. 2016: Internet. 5. O'Neal, Validation of twelve chemical spot tests for the detection of drugs of abuse. Forensic Science International, 2000. 109: p. 189-201. 6. Kelly, The Non-Specificity of the Duquenois-Levine Field Test for Marijuana. The Open Forensic Science Journal, 2012. 5: p. 4-8. 7. Jacobs, Detection of the Duquenois-Levine Chromophore in a Marijuana Sample. Forensic Science International, 2014. 239: p. 1-5. 8. Rubiano, The effect of benzene ring substituents on the mechanism of Duquenois Levine (DL) test for cannabinoid detection. Journal of Chemical and Pharmaceutical Research,, 2014. 6(6): p. 1261-1264. 9. Zaglul, J., An Analysis of Froehde’s and Duquenois-Levine Colorimetric Tests, in Biological Sciences. 2013, Wilkes Honors College of Florida Atlantic University. p. 23. 10. Kovar, Chemistry and reaction mechanisms of rapid tests for drugs of abuse and precursors chemicals, in UN Scientific and Technical Notes. 1989, The United Nations. p. 1-19. 11. Kelly, J., False Positives Equal False Justice. 2008: p. 1-44. Frank Talk - The Science of Cannabis: Powdery Mildew Mitigation and Strain Triage System7/19/2016
https://www.facebook.com/events/486473718220320/
The Endocannabinoid System: Truths, Half-Truths, and Unknowns Scientists frequently speak in caveats because there are few absolute truths in science; what is "true" (relevant) in one set of circumstance may not be true at all in a different context. The endocannabinoid system mediates a range of physiological responses that seem paradoxical. Activation of the CB2 receptor by THC, for example, suppresses immune cells that cause Crohn's Disease, but does not cause an overall state of immune-compromise. Cannabidiol (CBD), despite its structural similarity to THC, is not psychoactive, does not bind either of the two known endocannabinoid receptors (CB1 and CB2), yet somehow mimics THC's anti-inflammatory properties. This month's Frank Talk will explore some of what is currently known about the endocannabinoid system, attempt to dispel a few misunderstandings, and discuss promising new avenues of research. Cost: Free Date: Tuesday, May 31st 6:00-9:00 Location: Fort Greene Bar (Globeville) 321 E 45th Ave, Denver, Colorado 80216 Join Frank Conrad, Lab Director of Colorado Green Lab for The SCIENCE! of Cannabis. Happy hour is from 5:00-6:30 with half off wine bottles and taps all night for all you lovers of SCIENCE! Frank will talk from 6:00-6:30 and then open the floor for questions. The image below is of an insect called a thrip.
Thrips species feed on a wide array of plants and other insects by first puncturing them with their mandibles (clearly visible in image), then drawing out the nutrients through a tube-like structure in the mandibles called a stylet. Some species of thrips are predatory, and may be deliberately released in cannabis grows to combat spider mites and other parasites, but the majority of thrips are phytophagous ("plant-eating") insects that feed on chloroplasts from epidermal and mesophyll cells on the surface of the plant. Wikipedia article for general information on thrips: https://en.wikipedia.org/wiki/Thrips https://www.facebook.com/events/194673184226980/permalink/207296722964626/
The Chemicals Between Us: Imidacloprid Part 1 of a series examining the implications of pesticide contamination in cannabis products. This week's Frank Talk will compare EPA-established tolerance levels of imidacloprid in other commodities versus levels found in cannabis flower and concentrates, imidacloprid's mechanism of action, and discussion of potential health concerns for cannabis consumers. Cost: Free Date: Tuesday, March 15th 5:30-8:30 Location: Fort Greene Bar (Globeville) 321 E 45th Ave, Denver, Colorado 80216 Join Frank Conrad, Lab Director of Colorado Green Lab for The SCIENCE! of Cannabis. Happy hour is from 5:00-6:30 with half off wine bottles and taps all night for all you lovers of SCIENCE! Frank will talk from 6:30-7:00 and then open the floor for questions. 24-March-2016 Update:
The infection may have been caused by not one, but two fungal organisms; an epiphytic form of powdery mildew (possibly Podosphaera macularis) and endophytic false powdery mildew (probably Trichothecium roseum). More details and full update later. Powdery Mildew Disease of Cannabis Colorado Green Lab, in collaboration with Hempirical, is examining the life cycle, pathogenicity, and infection patterns in PM-resistant and PM-susceptible strains of cannabis. Powdery mildew may cause substantial economic loss directly by negatively impacting plant growth and flower quality, and indirectly if treated plants are rendered unsuitable for human consumption by accumulation of unsafe levels of fungicide residue. This is particular relevant where microorganisms have developed high levels of resistance to common fungicides (Demethylation Inhibitors such as myclobutanil), and cultivators apply ever-increasing amounts of fungicide just to keep the mildew in check. By elucidating the underlying cellular and genetic mechanisms behind cannabis powdery mildew susceptibility (and resistance), we aim to develop more effective means of combating the pathogen and reduce cultivation losses. Background Powdery mildew is not caused by a single organism, but by many different species of fungi. Most PM-species are highly specific to their preferred host and have evolved to attack a limited range of somewhat similar plants (e.g. cucumber mildew fungi cannot cause oak tree mildew, but can cause disease in squash plants). All PM-species are biotrophic, meaning they require living host tissue to grow and reproduce, and most share common infection mechanisms, with similar life cycles. Powdery mildew fungi are predominantly epiphytic ("epi" = "upon, near to", "phyt" = "plant") and grow on and around the surface of the plant, with some variants that also produce full or partially endophytic ("endo" = "within") mycelium and hyphae inside the infected plant tissue. Although the underlying mechanism behind resistance/susceptibility remains unclear, certain strains of cannabis are highly susceptible or highly resistant to infection by fungal organisms. The susceptible strain examined here is Blueberry, an Afghan indica hybrid (20% sativa / 80% indica). Infection of Susceptible Afghan Indica Powdery mildew infection of cannabis occurs on the adaxial (upper) surface of the leaf, which is coated with fine hairs (A-D) that may facilitate spore adhesion and subsequent infection. Infection occurs in stages. Spores landing on the upper surface of the leaf (E) germinate and form appressorium (F), flattened appendages that facilitate penetration. Appressorium form one or more haustorium (G), which are specialized appendages that punch a hole in the leaf surface, then act as feeding tubes that withdraw nutrients from the living plant. In PM-resistant cannabis strains, stomata automatically close soon after spore attachment (not shown), possibly to shut down photosynthesis by the affected tissue and starve out the infecting fungus. In susceptible cannabis strains (e.g. "Blueberry"), stomata are locked open directly below the site of the infection (H), allowing the infected tissue to continue photosynthesis and direct nutrients to the invading fungal haustorium. Following successful penetration of the leaf surface, filamentous mycelial growth invades the plant tissue, spreading throughout the interior of the leaf (I) and emerging from the lower (abaxial) leaf surfaces through stomata (J). Mycelium also forms on the exterior of the upper leaf surface and forms spore-bearing conidiophores (K), which release spores into the air and continue the cycle of infection. Surprise Guest Say hello to my little friend! Russet mite (probably...I'm not an entomologist) was found unexpectedly on the surface of a PM-infected leaf I was examining. Microscopy Notes Leaves were destained with Visikol (http://visikol.com/) to improve visualization. For added contrast of fungal and plant structures, some samples (those with reddish hue) were stained with modified Lugol's reagent, followed by dilute sulphuric acid. Thanks and credit to Koroch et al for the useful staining technique described in the literature referenced below. 1. Appl Plant Sci. 2013 Jul 5;1(7). Rapid staining method to detect and identify downy mildew (Peronospora belbahrii) in basil. Koroch AR, Villani TS, Pyne RM, Simon JE. Powdery mildew organisms are highly specific for their host, and P. fraxini, the causal agent shown here, doesn't infect cannabis. We used lilac leaves and associated PM as control samples to validate some new sample preparation, staining and microscopy techniques. Powdery mildew genera are grouped into five tribes: Phyllactineae, Erysipheae, Blumeriae, Golovinomyceteae, and Cystotheceae. The tribes can be identified by several criteria, including the type of host plant (Ex. Phyllactineae infects trees and shrubs), and by microscopic examination of their chasmothecium and appendage morphology, as well as shape and structure of spores. Cindy and I collected took Pee Em from a highly infected Lilac bush and brought it back to the lab. The dirty little %$#@ fungus wouldn't talk tho, and that's when things got a bit rough. We made the parasite spill its guts, all of them. Host plant (Lilac) and appendage morphology ("spines") positively identifies genera as Phyllactinia. Organism is most probably P. fraxini, which is known to infect Lilac leaves. Powdery mildew disease of cannabis in upcoming blog posts.
We've been hilariously busy with various projects....and then six months flew by without a blog post. Ouch. Blog content will be updated weekly from this point forward, documenting some of Colorado Green Lab's projects and interest areas. Upcoming posts:
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